Electronic cigarette (EC) use has increased dramatically in recent years, especially among never smokers. We recently completed a survey of EC users in Baltimore and found that a surprising percentage of EC users are non-smoking young adults. The oral cavity is host to one of the most complex microbial floras in the human body, and disturbance in this equilibrium can lead to overrepresentation of pathogenic species. Alterations in oral host defense, including reductions in salivary flow and secretion of antimicrobial peptides and cytokines by salivary glands, epithelial cells, and immune cells, may create a hospitable environment for pathogenic bacteria, leading to oral diseases. Our recent report indicates that EC exposure increases oxidative stress and inflammation, hampers the innate immune response, and increases susceptibility to pulmonary bacterial infections in a mouse model, indicating a high potential for adverse effects to oral host defense. EC exposure also causes inflammation of the salivary gland ducts in this model. However, a mouse model for studying effects on oral health is not representative of human exposure because mice are obligate nose breathers. EC vapor exposure in cell culture is also not representative of human exposure. Furthermore, the development of oral disease requires many years of exposure. Thus, cross-sectional and longitudinal alterations to the host oral defense and microbiome, which are early markers of risk, as a result of chronic exposure to ECs in young adults needs to be deciphered. We hypothesize that chronic EC use in young adults impairs the oral host defense by increasing oxidative stress and cytokines, and reducing salivary flow, antimicrobial peptides and epithelial defensins, leading to oral microbial dysbiosis. Specific Aim 1. Community structure and functional characterization of the oral microbiome in young adult EC users. Using samples from a cohort of young adult EC users and controls (age, 18-34 years; N=100/group), structural and functional characterization of the oral microbiome will be performed by 16S rRNA sequencing and metatranscriptomics for (a) cross sectional comparison of EC users with non-users and (b) a longitudinal follow up of EC users at 0, 1, and 2 years. Data from this Specific Aim will demonstrate functional differences in oral microbiomes of EC users that correlate with mechanistic biomarkers of oral host defense, which may increase the risk for oral disease. Specific Aim 2. Mechanistic evaluation of oral host defense in EC users. Along with the microbiome assessment in Specific Aim 1, oral host defense will be assessed via (1) salivary gland function, including salivary flow rate and salivary gland-specific anti-microbial peptides; () epithelial/immune cell antimicrobial defenses, including secretion of anti-microbial peptides, cytokines, and oxidative stress; and (3) biomarkers of inflammation and oral health in saliva and buccal epithelial cells. The integrative analysis of Specific Aim 1 and 2 will fill a critical gap n our understanding of alterations in host oral defense and a potential microbial shift in EC users, which may predispose them to oral diseases; thus, providing the evidence-based foundation for future public health policies on EC use.